scholarly journals Molecular detection of all 34 distinct O-antigen forms of Shigella

2009 ◽  
Vol 58 (1) ◽  
pp. 69-81 ◽  
Author(s):  
Yayue Li ◽  
Boyang Cao ◽  
Bin Liu ◽  
Dan Liu ◽  
Qili Gao ◽  
...  
Keyword(s):  
Shigella Flexneri ◽  
Bacterial Species ◽  
Milk Powder ◽  
Shigella Dysenteriae ◽  
Detection Sensitivity ◽  
Clinical Samples ◽  
Shigella Boydii ◽  
O Antigen ◽  
Probe Concentration ◽  
O Antigens

Shigella is the cause of shigellosis or bacillary dysentery, the occurrence of which is estimated to be 165 million cases per year worldwide, resulting in 1.1 million deaths. Rapid and reliable assays for detecting and identifying Shigella in food, environmental and clinical samples are therefore necessary. Shigella species are traditionally identified by their O antigens. This study developed a DNA microarray targeting O-serotype-specific genes to detect all 34 distinct O-antigen forms of Shigella, including Shigella boydii types 1–18, Shigella dysenteriae types 1–13, Shigella flexneri types 1–5 and 6, and Shigella sonnei. A total of 282 strains were used to test the specificity of the microarray, including 186 Shigella and Escherichia coli representative strains, 86 Shigella clinical isolates and ten strains of other bacterial species that are commonly isolated from food or clinical stool specimens. The oligonucleotide probes were printed on the microarray in concentrations from 1 to 100 μM, and 10 μM proved to be the optimal probe concentration. The detection sensitivity for each serotype was 50 ng genomic DNA or 1 c.f.u. in 25 g milk powder sample following a 6 h enrichment in broth. The microarray is specific, sensitive and reproducible, and, to our knowledge, is the first report of a microarray for serotyping all O-antigen forms of Shigella.

scholarly journals Sequence Analysis of Four Shigella boydii O-Antigen Loci: Implication for Escherichia coli andShigella Relationships

2001 ◽  
Vol 69 (11) ◽  
pp. 6923-6930 ◽  
Author(s):  
Lei Wang ◽  
Wenjia Qu ◽  
Peter R. Reeves
Keyword(s):  
Escherichia Coli ◽  
Gene Clusters ◽  
Shigella Dysenteriae ◽  
E Coli ◽  
Shigella Boydii ◽  
Antigen Gene ◽  
O Antigen ◽  
Atypical Features ◽  
Serotype 5 ◽  
O Antigens

ABSTRACT Shigella strains are in reality clones ofEscherichia coli and are believed to have emerged relatively recently (G. M. Pupo, R. Lan, and P. R. Reeves, Proc. Natl. Acad. Sci. USA 97:10567–10572, 2000). There are 33 O-antigen forms in these Shigella clones, of which 12 are identical to O antigens of other E. coli strains. We sequenced O-antigen gene clusters from Shigella boydiiserotypes 4, 5, 6, and 9 and also studied the O53- and O79-antigen gene clusters of E. coli, encoding O antigens identical to those of S. boydii serotype 4 and S. boydii serotype 5, respectively. In both cases the S. boydii and E. coli O-antigen gene clusters have the same genes and organization. The clusters of both S. boydii 6 and S. boydii 9 O antigens have atypical features, with a functional insertion sequence and a wzx gene located in the orientation opposite to that of all other genes in S. boydii serotype 9 and an rmlC gene located away from other rml genes in S. boydii serotype 6. Sequences of O-antigen gene clusters from another threeShigella clones have been published, and two of them also have abnormal structures, with either the entire cluster or one gene being located on a plasmid in Shigella sonnei orShigella dysenteriae, respectively. It appears that a high proportion of clusters coding for O antigens specific toShigella clones have atypical features, perhaps indicating recent formation of these gene clusters.

scholarly journals Stable Chromosomal Expression of Shigella flexneri 2a and 3a O-Antigens in the Live Salmonella Oral Vaccine Vector Ty21a

2017 ◽  
Vol 24 (12) ◽  
Author(s):  
Madushini N. Dharmasena ◽  
Manuel Osorio ◽  
Kazuyo Takeda ◽  
Scott Stibitz ◽  
Dennis J. Kopecko
Keyword(s):  
Oral Vaccine ◽  
Shigella Flexneri ◽  
Serum Antibody ◽  
Vaccine Vector ◽  
Shigella Dysenteriae ◽  
Antigenic Determinants ◽  
Content Type ◽  
O Antigen ◽  
Shigella Flexneri 2A ◽  
O Antigens

ABSTRACT We have been exploring the use of the live attenuated Salmonella enterica serovar Typhi Ty21a vaccine strain as a versatile oral vaccine vector for the expression and delivery of multiple foreign antigens, including Shigella O-antigens. In this study, we separately cloned genes necessary for the biosynthesis of the Shigella flexneri serotype 2a and 3a O-antigens, which have been shown to provide broad cross-protection to multiple disease-predominant S. flexneri serotypes. The cloned S. flexneri 2a rfb operon, along with bgt and gtrII, contained on the SfII bacteriophage, was sufficient in Ty21a to express the heterologous S. flexneri 2a O-antigen containing the 3,4 antigenic determinants. Further, this rfb operon, along with gtrA, gtrB, and gtrX contained on the Sfx bacteriophage and oac contained on the Sf6 bacteriophage, was sufficient to express S. flexneri 3a O-antigen containing the 6, 7, and 8 antigenic determinants. Ty21a, with these plasmid-carried or chromosomally inserted genes, demonstrated simultaneous and stable expression of homologous S. Typhi O-antigen plus the heterologous S. flexneri O-antigen. Candidate Ty21a vaccine strains expressing heterologous S. flexneri 2a or 3a lipopolysaccharide (LPS) elicited significant serum antibody responses against both homologous S. Typhi and heterologous Shigella LPS and protected mice against virulent S. flexneri 2a or 3a challenges. These new S. flexneri 2a and 3a O-antigen-expressing Ty21a vaccine strains, together with our previously constructed Ty21a strains expressing Shigella sonnei or Shigella dysenteriae 1 O-antigens, have the potential to be used together for simultaneous protection against the predominant causes of shigellosis worldwide as well as against typhoid fever.

scholarly journals Molecular Modeling of the Shigella flexneri Serogroup 3 and 5 O-Antigens and Conformational Relationships for a Vaccine Containing Serotypes 2a and 3a

Vaccines ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 643
Author(s):  
Jason Hlozek ◽  
Sara Owen ◽  
Neil Ravenscroft ◽  
Michelle M. Kuttel
Keyword(s):  
Diarrheal Disease ◽  
Shigella Flexneri ◽  
Quadrivalent Vaccine ◽  
Slight Effect ◽  
Work Support ◽  
O Antigen ◽  
The Common ◽  
O Antigens ◽  
Chain Conformations ◽  
Shared Epitopes

The pathogenic bacterium Shigella flexneri is a leading global cause of diarrheal disease. The O-antigen is the primary vaccine target and distinguishes the 30 serotypes reported. Except for serotype 6, all S. flexneri serotypes have a common backbone repeating unit (serotype Y), with variations in substitution creating the various serotypes. A quadrivalent vaccine containing serotypes 2a and 3a (as well as 6 and Shigella sonnei) is proposed to provide broad protection against non-vaccine S. flexneri serotypes through shared epitopes and conformations. Here we model the O-antigen (O-Ag) conformations of serogroups 3 and 5: a continuation of our ongoing systematic study of the S. flexneri O-antigens that began with serogroup 2. Our simulations show that S. flexneri serogroups 2, 3, and 5 all have flexible O-Ags, with substitutions of the backbone altering the chain conformations in different ways. Our analysis suggests three general heuristics for the effects of substitution on the Shigella O-Ag conformations: (1) substitution on rhamnose C reduces the extension of the O-Ag chain; (2) substitution at O-3 of rhamnose A restricts the O-Ags to predominantly helical conformations, (3) substitution at O-3 of rhamnose B has only a slight effect on conformation. The common O-Ag conformations across serotypes identified in this work support the assumption that a quadrivalent vaccine containing serotypes 2a and 3a could provide coverage against S. flexneri serotype 3b and serogroup 5.

scholarly journals Bacteriophage-encoded glucosyltransferase GtrII of Shigella flexneri: membrane topology and identification of critical residues

2005 ◽  
Vol 389 (1) ◽  
pp. 137-143 ◽  
Author(s):  
Adele M. LEHANE ◽  
Haralambos KORRES ◽  
Naresh K. VERMA
Keyword(s):  
Sequence Similarity ◽  
Repeat Unit ◽  
Shigella Flexneri ◽  
Membrane Topology ◽  
Transmembrane Helices ◽  
O Antigen ◽  
Repeat Units ◽  
Related Group ◽  
Critical Residues ◽  
O Antigens

The Shigella flexneri serotypes differ in the nature of their O-antigens. The addition of glucosyl or O-acetyl groups to the common backbone repeat units gives rise to the different serotypes. GtrII glucosylates rhamnose III of the O-antigen repeat unit, thus converting serotype Y (which has no modifications to the basic O-antigen repeat unit) into serotype 2a, the most prevalent serotype. In the present study, the topology of GtrII has been determined. GtrII has nine transmembrane helices, a re-entrant loop and three large periplasmic regions. Four critical residues (Glu40, Phe414, Cys435 and Lys478) were identified in two of the periplasmic regions. Despite the lack of sequence similarity between GtrII and the Gtrs from other serotypes, three of the critical residues identified are conserved in the remaining Gtrs. This is consistent with some degree of mechanistic conservation in this functionally related group of proteins.

Molecular Strategies for the Detection, Identification, and Differentiation between Enteroinvasive Escherichia coli and Shigella spp.

2005 ◽  
Vol 68 (2) ◽  
pp. 239-245 ◽  
Author(s):  
CESAR I. BIN KINGOMBE ◽  
MARIA-LUCIA CERQUEIRA-CAMPOS ◽  
JEFFREY M. FARBER
Keyword(s):  
Escherichia Coli ◽  
Multiplex Pcr ◽  
Shigella Flexneri ◽  
Shigella Dysenteriae ◽  
Shigella Boydii ◽  
Epidemiological Tool ◽  
Pcr Rflp ◽  
Shigella Spp

A strategy for the detection, identification, and differentiation of enteroinvasive Escherichia coli (EIEC) and Shigella spp. has been developed. The strategy includes (i) a multiplex PCR for the amplification of two virulence genes, i.e., iuc (222 bp) and ipaH (629 bp); (ii) amplification of the ial gene (a 1,038-bp amplicon) located within a large plasmid; and (iii) restriction fragment length polymorphism (RFLP) of the ial gene amplicon. The multiplex PCR provided three patterns. Pattern 1 (iuc−/ipaH+) was found in 10 (67%) of 15 EIEC strains tested, pattern 2 (iuc+/ipaH−) in only 2 (4.4%) of 46 non-EIEC isolates, whereas pattern 3 (iuc+/ipaH+) was observed in all Shigella spp. and also in 5 (33%) of 15 EIEC strains tested. The pattern 3 EIEC strains were all positive for the ial gene. The PCR-RFLP of the ial gene amplicon using the endonuclease AclI was used to differentiate Shigella spp. from the EIEC strains that belonged to pattern 3. The ial gene was present in 21 (38%) of 56 and 6 (40%) of 15 Shigella spp. and EIEC strains tested, respectively. The PCR-RFLP of the ial gene amplicon divided the strains in two types. Type 1 did not contain the restriction enzyme site and was found in 6 (100%) of 6 EIEC strains, 4 (80%) of 5 Shigella boydii, and 4 (100%) of 4 Shigella dysenteriae strains tested. Type 2, which gave two fragments of 286 and 752 bp, was observed in 5 (83%) of 6 Shigella flexneri strains and 6 (100%) of 6 Shigella sonnei strains. Detection, identification, and differentiation of Shigella spp. and EIEC were achieved by analyses of the PCR patterns and RFLP types. To our knowledge, this is the first study to demonstrate a simple and rapid method for detecting, identifying, and differentiating, at the molecular level, Shigella spp. and EIEC strains. This method will have tremendous utility as an epidemiological tool and in helping to develop policies, risk assessments, and national and international methods for Shigella spp.

scholarly journals Development of a DNA Microarray Method for Detection and Identification of All 15 Distinct O-Antigen Forms of Legionella pneumophila

2013 ◽  
Vol 79 (21) ◽  
pp. 6647-6654 ◽  
Author(s):  
Boyang Cao ◽  
Fangfang Yao ◽  
Xiangqian Liu ◽  
Lu Feng ◽  
Lei Wang
Keyword(s):  
Dna Microarray ◽  
Legionella Pneumophila ◽  
Detection Sensitivity ◽  
Immunological Methods ◽  
Clinical Samples ◽  
Content Type ◽  
O Antigen ◽  
Detection And Identification ◽  
Legionnaires Disease ◽  
Reference Strains

ABSTRACTLegionellais ubiquitous in many environments. At least 50 species and 70 serogroups of the Gram-negative bacterium have been identified. Of the 50 species, 20 are pathogenic, andLegionella pneumophilais responsible for the great majority (approximately 90%) of the Legionnaires' disease cases that occur. Furthermore, of the 15L. pneumophilaserogroups identified, O1 alone causes more than 84% of the Legionnaires' disease cases that occur worldwide. Rapid and reliable assays for the detection and identification ofL. pneumophilain water, environmental, and clinical samples are in great demand.L. pneumophilabacteria are traditionally identified by their O antigens by immunological methods. We have recently developed an O serogroup-specific DNA microarray for the detection of all 15 distinct O-antigen forms ofL. pneumophila, including serogroups O1 to O15. A total of 35 strains were used to verify the specificity of the microarray, including 15L. pneumophilaO-antigen standard reference strains and sevenL. pneumophilaclinical isolates as target strains, seven reference strains of other non-pneumophila Legionellaspecies as closely related strains, and six non-Legionellabacterial species as nonrelated strains. The detection sensitivity was 1 ng of genomic DNA or 0.4 CFU/ml in water samples with filter enrichment and plate culturing. This study demonstrated that the microarray allows specific, sensitive, and reproducible detection ofL. pneumophilaserogroups. To the best of our knowledge, this is the first report of a microarray serotyping method for all 15 distinct O-antigen forms ofL. pneumophila.

INTERSUBGROUP AND INTRASUBGROUP ANTIGENIC RELATIONSHIPS WITHIN THE GENUS SHIGELLA: SUBGROUP A

1961 ◽  
Vol 7 (3) ◽  
pp. 303-308
Author(s):  
W. H. Ewing ◽  
Jane G. Johnson
Keyword(s):  
Shigella Dysenteriae ◽  
Shigella Boydii ◽  
O Antigens ◽  
Antigenic Relationships

Intersubgroup antigenic relationships between the O antigens of cultures of Shigella dysenteriae 2 and 10 (subgroup A) and those of strains of Shigella boydii 1 (subgroup C) and between the O antigens of cultures of S. dysenteriae 8 and those strains of S. boydii 15 were reported. Also, several previously reported intra- and inter-subgroup relationships were confirmed and further elucidated. The relationships were important in definitive typing of the bacteria.

scholarly journals Impact of insertion sequences on convergent evolution ofShigellaspecies

2019 ◽  
Author(s):  
Jane Hawkey ◽  
Jonathan M. Monk ◽  
Helen Billman-Jacobe ◽  
Bernhard Palsson ◽  
Kathryn E. Holt
Keyword(s):  
Evolutionary Dynamics ◽  
Shigella Flexneri ◽  
Bacterial Species ◽  
Genomic Variation ◽  
Shigella Dysenteriae ◽  
Insertion Sequences ◽  
Single Nucleotide Variants ◽  
Genomic Studies ◽  
Metabolic Reduction ◽  
Genome Scale

AbstractShigellaspecies are specialised lineages ofEscherichia colithat have converged to become human-adapted and cause dysentery by invading human gut epithelial cells. Most studies ofShigellaevolution have been restricted to comparisons of single representatives of each species; and population genomic studies of individualShigellaspecies have focused on genomic variation caused by single nucleotide variants and ignored the contribution of insertion sequences (IS) which are highly prevalent inShigellagenomes. Here, we investigate the distribution and evolutionary dynamics of IS within populations ofShigella dysenteriaeSd1,Shigella sonneiandShigella flexneri. We find that five IS (IS1, IS2, IS4, IS600and IS911) have undergone expansion in allShigellaspecies, creating substantial strain-to-strain variation within each population and contributing to convergent patterns of functional gene loss within and between species. We find that IS expansion and genome degradation are most advanced inS. dysenteriaeand least advanced inS. sonnei; and using genome-scale models of metabolism we show thatShigellaspecies display convergent loss of coreE. colimetabolic capabilities, withS. sonneiandS. flexnerifollowing a similar trajectory of metabolic streamlining to that ofS. dysenteriae. This study highlights the importance of IS to the evolution ofShigellaand provides a framework for the investigation of IS dynamics and metabolic reduction in other bacterial species.

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